Fidaxomicin jams Mycobacterium tuberculosis RNA polymerase motions needed for initiation via RbpA contacts

ELife
Hande BoyaciElizabeth A Campbell

Abstract

Fidaxomicin (Fdx) is an antimicrobial RNA polymerase (RNAP) inhibitor highly effective against Mycobacterium tuberculosis RNAP in vitro, but clinical use of Fdx is limited to treating Clostridium difficile intestinal infections due to poor absorption. To identify the structural determinants of Fdx binding to RNAP, we determined the 3.4 Å cryo-electron microscopy structure of a complete M. tuberculosis RNAP holoenzyme in complex with Fdx. We find that the actinobacteria general transcription factor RbpA contacts fidaxomycin, explaining its strong effect on M. tuberculosis. Additional structures define conformational states of M. tuberculosis RNAP between the free apo-holoenzyme and the promoter-engaged open complex ready for transcription. The results establish that Fdx acts like a doorstop to jam the enzyme in an open state, preventing the motions necessary to secure promoter DNA in the active site. Our results provide a structural platform to guide development of anti-tuberculosis antimicrobials based on the Fdx binding pocket.

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Citations

Nov 28, 2018·Journal of Bacteriology·Tomáš KoubaLibor Krásný
Sep 10, 2019·Nucleic Acids Research·Ranjit K PrajapatiGeorgiy A Belogurov
Dec 28, 2019·Transcription·Hande BoyaciElizabeth A Campbell
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Dec 20, 2020·Nature Communications·Tomáš KoubaLibor Krásný
Jan 1, 2021·The Journal of Biological Chemistry·Konstantin Brodolin, Zakia Morichaud
Oct 31, 2020·Nature Reviews. Microbiology·James ChenElizabeth A Campbell
Nov 18, 2020·Proceedings of the National Academy of Sciences of the United States of America·Mirjana LilicElizabeth A Campbell
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Jan 12, 2021·The Journal of Biological Chemistry·Konstantin Brodolin, Zakia Morichaud
Apr 23, 2021·Proceedings of the National Academy of Sciences of the United States of America·Ilona Christy UnartaXuhui Huang
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Apr 21, 2018·Journal of Natural Products·Haibo ZhangChangsheng Zhang

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Methods Mentioned

BETA
FRET
size exclusion chromatography
electrophoresis

Software Mentioned

Coot
MotionCor2
Chimera
cryoSPARC
Phenix
Gautomatch
EMAN2
SBGrid
Gctf
ImageJ

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